ABSTRACT The principle strategy to prevent seasonal influenza infection is immunization to generate virus neutralizing antibodies. The magnitude and quality of vaccine-induced neutralizing antibody responses depend to a significant extent on the sum of diverse T cell responses activated through the presentation of vaccine or infection-derived sequences. Carefully shaped effector and regulatory T cell responses can help generate protective humoral immunity, but an imbalance favoring regulatory mechanisms may impede antigen-specific B cell responses and limit the development of high affinity antibodies, resulting in poor vaccine efficacy. While much has been learned about influenza vaccine-specific effector T cell responses, most studies have largely overlooked mechanisms that diminish vaccine efficacy. We developed computational methods for identification of potential regulatory T cell-inducing epitopes in pathogen sequences that are highly cross-conserved with human protein sequences. Based on our preliminary studies showing that Treg activation and function is linked to viral T cell epitopes bearing homology to human sequences, we hypothesize that human-like epitopes in seasonal influenza virus stimulate suppressive mechanisms that limit vaccine efficacy. To test this hypothesis, we propose to conduct immune monitoring studies using an existing cell and serum bank from subjects immunized with the trivalent inactivated virus influenza vaccine over four consecutive years, beginning with the 2008-2009 influenza season. This unique cohort, spanning the pre- and post-2009 H1N1 pandemic eras, affords an opportunity to ask significant questions about changes in humoral and cellular immune responses to H1N1 and H3N2 vaccine strains within each season and from season to season. We will investigate associations between hemagglutination inhibition titers and frequencies of HA-specific CD4+ Tregs and circulating regulatory follicular T cells (cTfr). Additionally, we will evaluate the frequency and suppressive function of Treg and cTfr cells specific for human-like influenza A virus HA T cell epitopes in human vaccination. Identification of Treg-/Tfr-activating epitopes associated with limited antibody responses may begin to uncover a mechanism of immune-evasion underappreciated in seasonal influenza vaccination. Moreover, it may spur novel vaccine design strategies that increase the efficacy of influenza vaccines and vaccines in general.